1. Field of the Invention
This invention relates to a method for extracting a biosubstance from the root of a hair. More specifically, the present invention is concerned with a method for extracting a biosubstance such as RNA, DNA, protein or element from the root of a hair that requires pulling force of at least a predetermined reference value to pull it out and also with a hair sampling device useful in the method.
2. Description of the Related Art
These days, diverse gene expression analyses led by DNA microarrays are widely used to elucidate the molecular mechanisms of various diseases and also to search disease markers and drug targets.
For a DNA microarray, RNA is firstly extracted from a sample tissue or sample cells, and cDNA is then synthesized with a reverse transcriptase while using the RNA as a template. Upon this synthesis, the cDNA is labeled with a phosphor such as Cy3 or Cy5, followed by hybridization of the labeled cDNA with probe DNA immobilized on the microarray. Based on fluorescence signals from the hybridized cDNA, an expressed gene is analyzed.
DNA microarrays in the past have been considered to require RNA in a substantial quantity (on the order of several micrograms) for the provision of the above-described labeled cDNA. Accordingly, targets of analyses were limited to relatively large tissue fragments and large quantities of cells, and analyses of small tissue fragments and small quantities of cells such as biopsy samples were difficult.
In recent years, however, high-sensitivity DNA microarrays have been developed to permit measurements of nanogram-level gene samples, thereby making it possible to also perform gene expression analyses on micro-volume samples such as small tissue fragments and small quantities of cells. It has also become increasingly possible to perform micro-volume sample analyses in gene polymorphism analyses, proteome analyses and the like.
These micro-volume sample analyses have attracted increasing interests as permitting easy sample collections at actual clinical sites and also enabling to accumulate samples and their analysis data and also to establish fast diagnosis methods.
In a gene polymorphism analysis, for example, it has heretofore been necessary to sample blood as much as 10 mL or so to collect leucocytes from the blood, to lyse leucocytes to extract DNA, to amplify the DNA by a PCR, and then to perform a gene polymorphism analysis. At present, however, systems have been put on the market to perform the extraction of DNA from a micro-volume sample such as a drop of blood obtained by slightly puncturing a fingertip or an extremely small piece of mucosa collected by scratching off the tunica mucosa oris, its amplification and even the determination of its genotype in a fully automated fashion.
Keeping in step with the increasing practice of the analysis of biosubstances such as RNA, DNA, proteins and elements from such micro-volume samples, hair is attracting interests as samples in addition to the above-mentioned blood drops and tunica mucosa oris. Hair can be noninvasively and simply collected or sampled without imposing heavy physical or mental pain or suffering on donors (patients). Hair is, therefore, expected to serve as a suitable source for the extraction of biosubstances toward the popularization of order-made medical care such as the diagnosis of drug sensitivity by the single-base polymorphism (SNP) analysis of genes.
As a method for the diagnosis of a disturbance in physical conditions, a method to detect an element contained in a hair root has been developed (see Japanese Patent Laid-open No. 2004-45133 referred to as Patent Document 1 hereinafter).
Further, Japanese Patent Laid-open No. 2005-192409 (referred to as Patent Document 2 hereinafter) discloses, as a method for the recovery of RNA from hair such as head hair, a method that freezes up a hair to extreme cold with liquefied nitrogen immediately after its pull-out, brings the frozen hair into contact with an RNA extracting reagent, and then subjects the resulting hair to vortex agitation processing to extract RNA.